Nitric oxide (NO) has been shown in vitro to play an important role in the regulation of oxygen consumption on the mitochondrial respiratory chain. Our preliminary data demonstrated that oxygen consumption in the postischemic in vivo mouse heart was suppressed and the suppression showed a reversible and an irreversible phase. Whether NO can regulate mitochondrial respiration in vivo and whether this regulation plays a protective role in the postischemic myocardium still remains an important unresolved question. In order to determine the in vivo regulatory role of NO on mitochondrial respiration, real-time myocardial tissue oxygen consumption and tissue injury need to be assessed. However, until recently it was impossible to measure these critical parameters in the beating heart due to the lack of in vivo techniques. Recently, electron paramagnetic resonance (EPR) oximetry and imaging techniques have been developed using a novel surface coil resonator (SCR) with automatic tuning control (ATC), automatic coupling control (ACC), and cardiac gating capability. By employing these techniques, we are able to perform oximetry and imaging measurements on the in vivo mouse heart to determine: (1) real-time variations in tissue oxygen tension and blood flow for the assessment of oxygen consumption during and after regional ischemia and reperfusiion; (2) regulation of oxygen consumption in the postischemic myocardium by eNOS-derived NO and its derivative peroxynitrite; (3) real-time alterations in postischemic myocardial injury and thereby determine the protective role of NO via the suppression of oxygen consumption. Overall, we will utilize the unique capabilities of in vivo EPR spectroscopy and imaging to provide direct and noninvasive measurement and imaging of myocardial tissue pO2, tissue oxygen consumption, redox status, and viability. Critical information will be obtained regarding the regulatory role of endothelium-derived NO on myocardial oxygen consumption, redox alterations, and postichemic myocardial injury and protection in cardiac physiology and disease.